Vaccine development efforts against human immunodeficiency virus type 1 (HIV-1) are hindered by the high mutation rate of the virus, lack of clear understanding of the nature of the protective immunity, and limitations of available animal models for direct testing of HIV antigens for protection. We tested protective efficacy of a highly conserved HIV envelope peptide cocktail vaccine in Indian origin rhesus macaques by taking advantage of the chimeric virus SHIV, that expresses HIV envelope and causes AIDS in this model thereby allowing for direct testing of HIV vaccines based on envelope sequences. The HIV envelope peptides for the vaccine cocktail were selected through a series of studies in multiple animal models and in vitro studies with cells from HIV-infected long-term non-progressors, for broadly cross reactive T cell responses in the context of multiple MHC haplotypes. Since the peptide cocktail vaccine selectively primes cell-mediated immunity (CMI) in the absence of anti-HIV antibody production, it is possible to test protective efficacy of antiviral TH and CTL responses in this model. Three separate vaccine studies were conducted employing a total of 30 macaques for delivering the peptide cocktail vaccine with Freund's adjuvant and/or autologous dendritic cells (DC). Macaques immunized with ex vivo peptide cocktail-pulsed DC exhibited efficient induction of CMI, and upon challenge with pathogenic strains of SHIV (SHIV[ku2] and SHIV[89.6P]) showed significant reduction in viral set point accompanied by undetectable virus in the blood of majority of vaccinated animals. Importantly, no rebounds were observed in the vaccinated monkeys, some followed for over three years. These results strongly support the vaccine potential of the HIV envelope peptide cocktail and the efficiency of DC loaded ex vivo with peptides for priming protective CMI responses. We hypothesize that FIt3-ligand (FL) treatment followed by delivering the vaccine cocktail in the presence of CpG-containing oligodeoxynucleotides (ODN) will be an effective strategy for targeting the vaccine to DC in vivo. Further we propose that this strategy would entail the HIV envelope peptide cocktail to be a viable vaccine approach applicable to humans. We obtained preliminary data supporting FL-mediated mobilization of multiple subsets of DC with immunogenic properties in murine and primate studies. We will test whether FL treatment of macaques followed by vaccination with the HIV envelope peptide cocktail in CpG-ODN would be effective in priming strong CMI and protection against intravenous challenge using SHIV[KU2]. Since the major route of HIV infection is the mucosal epithelium and is predominantly by macrophage-tropic HIV strains, we will also test the effectiveness of the peptide cocktail targeted to DC in vivo using FL-treatment and CpG-ODN followed by mucosal challenge using SHIV[SF162P], (expressing macrophage-tropic HIV envelope). We will also test a mucosal vaccination strategy for delivering the peptide cocktail using a novel mutant cholera toxin (CT2*) that we observed to be a strong adjuvant for priming systemic and mucosal immune responses to several peptide antigens, including the HIV vaccine peptides in mice.